Despite increased survivorship among patients, breast cancer remains the second leading cause of cancer death in women. The magnitude of this problem provides a strong impetus for studies that may lead to the development of new chemopreventative strategies and/or lifestyle changes that reduce cancer incidence. In this regard, several studies positively correlate obesity to the development of breast cancer. Current evidence and proposed mechanisms for this observation strongly implicate the involvement of the estrogen receptor (ER). Importantly, obesity is also highly associated with elevated cholesterol, and cholesterol itself is a risk factor for breast cancer. Furthermore, patients takin lipophilic inhibitors of HMG-CoA reductase (statins) demonstrate a decreased risk for breast cancer incidence and recurrence. The recent observation that 27- hydroxycholesterol (27HC) is produced in a stoichiometric manner from cholesterol, together with our data demonstrating that it exerts partial agonist activity on both the ERs and liver X receptors (LXRs), suggests a potential, heretofore unrecognized, mechanistic link between hypercholesterolemia and breast cancer incidence. Several key pieces of evidence form the foundation of the proposed project: (1) obesity, metabolic syndrome and high serum cholesterol are all associated with increased breast cancer incidence, (2) enrichment of tumor-associated macrophages, the primary producers of 27HC, in the tumor microenvironment is associated with a poor prognosis, and (3) elevation of 27HC levels in a murine model of spontaneous mammary cancer increases tumor growth and metastasis in vivo. Building on these findings we hypothesize that 27HC, through its actions on ER and LXR, is an important mechanistic link between obesity, hypercholesterolemia and increased incidence of breast cancer. My long-term goal is to develop an independent research program focused on defining how manipulation of cholesterol biology, pathologically or pharmacologically, impacts the initiation and progression of breast cancer. With the help of the highly qualified mentors and advisory committee members that I have selected, along with the exceptional research environment at Duke University School of Medicine, I am very well situated to address the important questions being asked in this proposal. At the conclusion of this project I expect to have (1) established mechanistic links between obesity, cholesterol and increased susceptibility, growth and metastasis of breast cancer, (2) evaluated the utility of targeting cholesterol and 27HC production, and relevant downstream signaling pathways in the treatment and prevention of breast cancer, and (3) defined the mechanisms by which 27HC increases metastasis. It is anticipated that this line of investigation will lead to the near-term development of new strategies to treat and prevent breast cancer. Furthermore, the results will likely provide additional support and rationale for the exploration of potential chemopreventative benefits of lower cholesterol diets and/or pharmacological inhibitors of HMG-CoA reductase or CYP27A1.